The goal of the proposed research is to elucidate the function and reactivity of hydroxylamine-cytochrome c554 oxidoreductase (HAO), a remarkably complex respiratory enzyme from the bacterium Nm. europaea. A series of novel pulsed-laser techniques will be used to generate postulated intermediates of HAO's catalytic pathway, and a combination of electrochemical and computational techniques will be used to investigate the properties of the enzyme's many redox-active centers. The net oxidation of ammonia to nitrite by oxygen is the only process whereby Nm. europaea can extract usable energy from the surroundings. HAO catalyzes a critical step in this process, the 4-electron oxidation of hydroxylamine to nitrite. Bacteriologically-mediated oxidation of ammonia to nitrite is an important public and environmental health issue in all parts of the country where agriculture is a significant part of the economy. Nitrite is a potential precursor for carcinogenic N-nitroso compounds, and for the potent greenhouse gas nitrous oxide. As much as 50% of ammonia fertilizer may be oxidized to nitrite by soil-inhabiting bacteria before it can reach its target crops, and because nitrite is highly soluble, it readily contaminates surrounding rivers and groundwater. Under carefully controlled conditions bacteriologically-mediated ammonia oxidation can also benefit public health. Mixtures of Nm. europaea, and complementary bacteria that reduce nitrite to nitrogen, are used in sewage treatment plants to eliminate excess ammonia from waste water. The research being proposed, which aims at understanding in detail how HAO works, may suggest ideas about how to manage ammonia oxidation in agricultural settings and sewage treatment, for maximum benefit to public and environmental health. The project may also provide important insights into the redox chemistry of physiologically important enzymatic metal-nitrogen species that will be of interest to the medical research community. [unreadable] [unreadable] [unreadable]